Microfluidic PCR has evolved since Wilding and coworkers first performed PCR in a chamber in a microchip device (Wilding, P., Shoffner, M. A., Kricka, L. J., PCR in a silicon microstructure, Clin. Chem., 1994, 40, 1815-1818). Northrup et al. described a device that coupled a PCR reactor and a capillary electrophoresis (CE) module fabricated on different substrates (Woolley, A. T., Hadley, D., Landre, P I, deMello A. J., Mathies, R. A., Northrup, M. A. Functional integration of PCR amplification and capillary electrophoresis in a microfabricated DNA analysis device, Anal. Chem., 1996, 68, 4081-4086). Later, Burns' group developed an integrated device which could perform PCR and gel-based electrophoresis (Burns, M. A., Johnson, B. N., Brahmasandra, S. N., Handique, K, Webster, J. R., Krishnan, M., Sammarco, T. S., Man, P. M., Jones, D., Heldsinger, D., Mastrangelo, C. H., Burke, D. T., An integrated nanoliter DNA analysis device, Science, 1998, 282, 484-487). Lagally from Mathies group and Koh from ACLARA Biosciences also demonstrated PCR-CE in integrated microfluidic devices (Lagally, E. T., Simpson, P. C., Mathies, R. A., Monolithic integrated microfluidic DNA amplification and capillary electrophoresis analysis system, Sensors and Actuators B, 2000, 63, 138-146; and Koh, C. G., Tan, W., Zhao, M., Ricco, A. J., Fan, Z. H., Integrating polymerase chain reaction, valving, and electrophoresis in a plastic device for bacterial detection, Anal. Chem., 2003, 75, 4591-4598). Hess et al. used a reactor to carry out PCR under high pressures to control nucleic acid hybridization (Hess, R. S., Laugharn, J. A. Jr., Green, D. J., Pressure-controlled nucleic acid hybridization, U.S. Pat. No. 6,753,169B2, Jun. 22, 2004).
The temperature needed to conduct PCR can reach up to 95° C., which is close to the boiling point of water. At such a high temperature, evaporation is severe and it can change the concentration in the reaction solution and lower the PCR efficiency. Bubbles can be generated inside the solution in the PCR chamber, generating pressure differences in microfluidic channels and pushing liquid out of intended regions; for example, a separation buffer can be moved out of a CE separation channel. Further, the valves, such as gel valves, wax valves, and hydrophobic material generally used in many of the microfluidic devices are not reusable, limiting the devices to the detection of PCR amplification only at the final phase or at the end-point of the PCR reaction.
Disposable PCR devices are desirable to avoid carryover and cross-contamination issues. Moreover, although valves can be used to prevent evaporation and liquid movement, incorporation of valves into microfluidic PCR devices will substantially increase the cost of fabrication. Use of valves in microfluidic PCR systems is also questionable because they tend to lose their functions once they are activated and, consequently, do not allow continuous or multiple sampling of products from the reaction chamber.
A device is presented with a manifold used to suppress or prevent evaporation, condensation, and unintended movement of liquid because of pressure differences, for example, in a microfluidic channel network during PCR cycles.